title: Combination of necroptosis and apoptosis inhibition enhances cardioprotection against myocardial ischemia-reperfusion injury.
abstract: OBJECTIVE
  Necroptosis has been proposed as a mode of cell death that is a caspase-independent programmed necrosis. We investigated whether necroptosis is involved in myocardial ischemia-reperfusion injury in isolated guinea pig hearts and, if so, whether simultaneous inhibition of necroptosis and apoptosis confers enhanced cardioprotection.
  
  
  METHODS
  Isolated perfused guinea pig hearts were subjected to 30 min ischemia and 4 h reperfusion (control = CTL, n = 8). Necrostatin-1 (necroptosis inhibitor, 10 μM), Z-VAD (apoptosis inhibitor, 0.1 μM) and both inhibitors were administered starting 5 min before ischemia and during the initial 30 min of reperfusion (Nec, Z-VAD, Nec + Z-VAD; n = 8 each). Contractile recovery was monitored by left ventricular developed (LVDP) and end-diastolic (LVEDP) pressure. Infarct size was determined by triphenyltetrazolium chloride staining. Tissue samples were obtained after 4 h reperfusion to determine expression of receptor-interacting protein 1 (RIP1) and activated caspase 3 by Western blot analysis.
  
  
  RESULTS
  After reperfusion, Nec + Z-VAD had higher LVDP and lower LVEDP compared with CTL. Infarct size was reduced in Nec and Z-VAD compared with CTL. Combination of necroptosis and apoptosis inhibition further reduced infarct size. Expression of activated caspase 3 was not increased in Z-VAD and Nec + Z-VAD compared with Nec and CTL. Expression of RIP1 was preserved in Z-VAD and Nec + Z-VAD compared with CTL, suggesting RIP1-mediated necrosis is involved in myocardial ischemia-reperfusion injury.
  
  
  CONCLUSIONS
  Necroptosis is involved in myocardial ischemia-reperfusion injury, and simultaneous inhibition of necroptosis and apoptosis enhances the cardioprotective effect. These findings may provide a novel, additive strategy for cardioprotection in acute myocardial infarction.
authors.affiliation: Department of Anesthesiology, Graduate School of Dentistry, Osaka Dental University, Osaka, Japan.
authors.forename: Shizuka
authors.initials: S
authors.lastname: Koshinuma
authors.forename: Masami
authors.initials: M
authors.lastname: Miyamae
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Kaneda
authors.forename: Junichiro
authors.initials: J
authors.lastname: Kotani
authors.forename: Vincent M
authors.initials: VM
authors.lastname: Figueredo
mesh_terms: D000590:Amino Acid Chloromethyl Ketones / Q000008:administration & dosage / Q000627:therapeutic use*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002316:Cardiotonic Agents / Q000008:administration & dosage / Q000627:therapeutic use*; D053148:Caspase 3 / Q000032:analysis; D004359:Drug Therapy, Combination; D006168:Guinea Pigs; D006321:Heart / Q000187:drug effects* / Q000503:physiopathology; D006439:Hemodynamics / Q000187:drug effects; D007093:Imidazoles / Q000008:administration & dosage / Q000627:therapeutic use*; D007211:Indoles / Q000008:administration & dosage / Q000627:therapeutic use*; D008297:Male; D015428:Myocardial Reperfusion Injury / Q000188:drug therapy* / Q000473:pathology / Q000503:physiopathology; D009206:Myocardium / Q000473:pathology*; D009336:Necrosis / Q000188:drug therapy / Q000473:pathology / Q000503:physiopathology
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D002316:Cardiotonic Agents; D007093:Imidazoles; D007211:Indoles; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C507699:necrostatin-1; D053148:Caspase 3